Effects of 2 water-soluble contrast media, meglumine iothalamate (Conray) and meglumine iocarmate (Dimer-X), on the electric activity of identifiable giant neurons of the African giant snail (Achatina fulica Ferussac)]

The influences of two water soluble contrast media, meglumine iothalamate and meglumine iocarmate, on the neuronal excitability and on the neuronal sensitivity to putative transmitters were examined in comparison with those of sucrose using two identifiable giant neurones of Achatina fulica Férussac (the TAN and the PON). A relatively low increase of osmotic pressure of the extracellular fluid, produced by the application of contrast media, reversed the Cl- dependent inhibition caused by a putative transmitter. The same increase of this osmotic pressure, however, did not influence the Cl- independent inhibition and the excitation of the neurone examined. The hyperpolarization of neuromembrane was caused by an increase of osmotic pressure of the extracellular fluid. Its relatively high increase was necessary to make spontaneous spike discharges disappear totally. All effects of the two contrast media, observed in this study, were due to the increase of osmotic pressure of the extracellular fluid ; no specific effect of the contrast media containing the iodine on the indicators used was observed.     

Measurement of Serine Acetyltransferase Activity in Crude Plant Extracts by a Coupled Assay System Using Cysteine Synthase

Serine acetyltransferase (SATase) (EC 2.3.1.30 [EC] ) catalyzes theformation of Oacetyl-L-serine (OAS) from L-serine in the presenceof acetyl-CoA. A novel assay method was developed for measuringthis enzyme activity in extracts from plant tissues. The assayconsists of a coupled system in which the OAS formed is convertedto cysteine by the addition of cysteine synthase (CSase) (EC4.2.99.8 [EC] ). Cysteine thus formed is determined colorimetricallyand serves as a measure for SATase activity. This method israpid, simple and sensitive, and can be readily adapted formeasurement of SATase activity in crude tissue extracts or homogenates. (Received January 14, 1987; Accepted April 27, 1987)     

Assessing the Patterns of Evolution in Anuran Vocal Sexual Signals

Many animal species have evolved signalling traits to mediate various intra-specific interactions. Signals are particularly important for inter-sexual selection, where females use male signalling traits to select mates. Female preferences are therefore a major selective force in the evolution of these male signals, and these preferences can facilitate rapid changes in these traits in an evolutionary timeframe. This introduction of high levels of variation in inter-sexual signals may overshadow any phylogenetic patterns present. Such shadowing effects, however, should be dependant on the characteristics of traits (e.g. morphological, physiological and behavioural). Using male advertisement calls from 72 species of anuran amphibians, we tested the levels of phylogenetic signal present for a variety of call features in relation to trait types, and for calls as whole units using phylogenetic principal components analysis. We found that most call features displayed some level of phylogenetic autocorrelation (or signal), with traits that are dependent on morphology having much stronger phylogenetic signals than those based on behaviour. In addition, when calls were analysed as whole units, closely related species were found to be similar to each other, indicating that phylogenetic patterns had not been cancelled out by selection via female preferences. We suggest that signal functions, such as indicating male quality (e.g. mediated by body size) to potential mates, may place constraints on the amount of variation that can be introduced by female preferences. More research, particularly studies on other taxa, will be required to elucidate whether the patterns found in anurans are general across the animal kingdom.     

Identification and purification of sulfotransferases for 20-hydroxysteroid from the larval fat body of a fleshfly,Sarcophaga peregrina

Sulfotransferase (ST) activity for 20-hydroxyecdysone (20E) was identified in a larval fat body lysate of the fleshfly, Sarcophaga peregrina, but not in the hemolymph. The activity was highly sensitive to 2,6-dichloro-4-nitrophenol (DCNP) (IC50=0.61 microM), a specific inhibitor of phenol ST (P-ST), but insensitive to triethylamine, a hydroxysteroid ST inhibitor. These results suggest that 20E-specific ST enzymes belong to the P-ST family, despite the fact that 20E is a hydroxysteroid. In addition to 20E ST activity, a relatively high level of 2-naphthol ST activity was detected in the fat body lysate. The ST activity for both substrates transiently decreased to the 50% of maximal levels, 6 hrs after induction of pupation. The ST enzymes were separated on a DEAE-cellulose column. The 20E-ST enzymes were eluted around 50 mM KCl as two separate peaks of close proximity and the P-ST was eluted at 0.1 M KCl. The 20E ST enzymes were further purified using 3'-phosphoadenosine 5'-phosphate (PAP)-agarose affinity column chromatography. Both of the eluted active fractions demonstrated 43-kDa proteins on SDS-polyacrylamide gel. Photoaffinity labeling with [35S]-3'-phosphoadenosine 5'-phosphosulfate (PAPS) showed 43-kDa bands in the fat body lysate, as well as in the purified fractions. These results suggest that the 43-kDa proteins catalyze 20E sulfation within the fat body of S. peregrina.     

Functional analysis of chick heparan sulfate 6‐O‐sulfotransferases in limb bud development

Heparan sulfate (HS) interacts with numerous growth factors, morphogens, receptors, and extracellular matrix proteins. Disruption of HS synthetic enzymes causes perturbation of growth factor signaling and malformation in vertebrate and invertebrate development. Our previous studies show that the O‐sulfation patterns of HS are essential for the specific binding of growth factors to HS chains, and that depletion of O‐sulfotransferases results in remarkable developmental defects in Drosophila, zebrafish, chick, and mouse. Here, we show that inhibition of chick HS‐6‐O‐sulfotransferases (HS6ST‐1 and HS6ST‐2) in the prospective limb region by RNA interference (RNAi) resulted in the truncation of limb buds and reduced Fgf‐8 and Fgf‐10 expressions in the apical ectodermal ridge and in the underlying mesenchyme, respectively. HS6ST‐2 RNAi resulted in a higher frequency of limb truncation and a more marked change in both Fgf‐8 and Fgf‐10 expressions than that achieved with HS6ST‐1 RNAi. HS6ST‐1 RNAi and HS6ST‐2 RNAi caused a significant but distinct reduction in the levels of different 6‐O‐sulfation in HS, possibly as a result of their different substrate specificities. Our data support a model where proper levels and patterns of 6‐O‐sulfation of HS play essential roles in chick limb bud development.     

New Rev-export inhibitor from Alpinia galanga and structure–activity relationship

Bioassay-guided separation by use of the fission yeast expressing NES of Rev, an HIV-1 viral regulatory protein, disclosed 1′-acetoxychavicol acetate (ACA, 1) as a new inhibitor for nuclear export of Rev from the roots of Alpinia galanga. Both analysis for mechanism of action with biotinylated probe (2) and several synthesized analogs established crucial portions in 1 for Rev-export inhibitory activity.     

Gastrointestinal flora of cotton rats

The gastrointestinal (GI) flora of cotton rats was examined. No lactobacilli were detected in any part of the GI tract. Anaerobes, including Peptococcaceae, Bacteroidaceae, bifidobacteria and eubacteria, were the predominant bacteria in the stomach, small intestine, caecum and faeces. Aerobes and facultative anaerobes, including Enterobacteriaceae and streptococci, were detected at low numbers and very low frequency of occurrence in all parts of the GI tract. Sixty-one isolates of bifidobacteria were recovered from the stomach, small intestine, caecum and faeces of cotton rats. They were identified as Bifidobacterium animalis, B. pseudolongum biovar a and b. The study showed that the GI flora of cotton rats seem to be very different from the GI flora in other rodents.     

Primary structure of rat insulin-like growth factor-I and its biological activities

Rat insulin-like growth factor-I (IGF-I), a serum polypeptide with growth promoting activity, was isolated from rat serum by a combination of acid/ethanol extraction, affinity chromatography, and a series of reversed phase high performance liquid chromatography, cation exchange, and reversed phase. All peptide fragments produced by chymotrypsin digestion of reduced and carboxymethylated rat IGF-I were amino acid sequenced and compared with the sequence of human IGF-I. Three out of 70 of the rat amino acid residues differed from those of human IGF-I as follows: Asp20----Pro, Ser35----Ile and Ala67----Thr. Purified rat IGF-I cross-reacted with polyclonal anti-human IGF-I antibody 75% as compared to human IGF-I, but it cross-reacted only 3% with monoclonal anti-human IGF-I antibody. Thus, it is possible to monitor the metabolic fate of human IGF-I, when injected into rats, without interference by endogenous rat IGF-I. Rat IGF-I showed 65% activity in the radioreceptor, 28.6% activity in the lipogenesis and 22.5% activity in the free fatty acid release inhibition assays as compared to human IGF-I on a protein quantity basis.     

Comparison of in vitro susceptibilities of Rickettsia prowazekii, R. rickettsii, R. sibirica and R. tsutsugamushi to antimicrobial agents

The in vitro activities of 16 antimicrobial agents against Rickettsia prowazekii (Breinl strain), R. rickettsii (Bitterroot strain), R. sibirica (ATCC No. VR151) and R. tsutsugamushi (Gilliam, Karp, Kato, Shimokoshi, Kawasaki and Kuroki strains) were determined by the cell culture method. Tetracycline, demethylchlortetracycline, doxycycline, minocycline, chloramphenicol, kitasamycin and rifampicin were generally effective (MIC, 0.005-0.78 micrograms/ml) to all strains tested. Quinolones such as norfloxacin, ciprofloxacin and ofloxacin were moderately active, but they were less active against R. tsutsugamushi than other rickettsial species. Penicillins and cephems showed low activity against most of the strains tested, but high concentrations of benzylpenicillin (MIC, 25-50 micrograms/ml) inhibited R. prowazekii, R. rickettsii and R. sibirica. These findings may be applicable for differentiation of species of genus Rickettsia.